Immunogenicity of synthetic HIV-1 gp120 V3-loop peptide-conjugate immunogens

A successful prophylactic human immunodeficiency virus type 1 (HIV-1) vaccine must elicit an immune response that will prevent establishment of the persistent viral infection. The only response shown to be effective in this regard is virus-neutralizing antibody directed against the viral gp120 hypervariable V3-loop region. Conjugate immunogens, containing cyclic peptides representing the V3 determinant covalently bound to a carrier protein, were capable of eliciting virus-neutralizing antibodies. The consistency of the response was related to peptide size. The smaller cyclic peptides, expressing relatively conserved sequences from the V3-loop apex, were poor inducers of neutralizing activity. In contrast, the largest cyclic peptides mediated neutralizing responses that were similar to those observed and previously reported for intact gp120 immunogens. A cyclic synthetic peptide expressing most of the prototypic HIV-1 MN variant V3 determinant warrants further study as a potentially effective vaccine immunogen.     

Sera from HIV-1 infected individuals in all stages of disease preferentially recognize the V3 loop of the prototypic macrophage-tropic glycoprotein gp120 ADA

The outer membrane glycoprotein gp120 and the transmembrane glycoprotein gp41 are predominant targets of the humoral immune response to infection by human immunodeficiency virus type 1. The third hypervariable region (V3 loop) is the principal neutralizing domain and is the primary target of neutralizing antibodies directed against the envelope proteins of HIV-1. The V3 loop is also the major determinant for HIV-1 cell-specific tropism. To further characterize the humoral immune response directed against the gp120 envelope proteins, we expressed two prototypic gp120 envelope proteins (LAI/HXB2 and ADA) and chimeric gp120 envelope proteins in stable transfected Drosophila melanogaster Schneider 2 cells. Sera from four infected adults over the course of infection [McNearney et al. (1992) Proc. natn. Acad. Sci. U.S.A. 89, p. 10,242] were assayed for reactivity with the respective envelope proteins. Sera obtained at early stages preferentially recognized the gp120 envelope protein ADA, whereas in later stages of infection the sera showed diminished reactivity with both gp120 LAI/HXB2 and gp120 ADA. Chimeric envelope proteins revealed that the humoral response was directed primarily against the V3 loop of gp120 ADA. Furthermore, 22 sera from HIV-1 infected individuals in different stages of the disease were tested. Reactivity of sera with the gp120 envelope protein ADA was seven-fold higher than with the gp120 envelope protein LAI/HXB2. Our results suggest that the humoral immune response is preferentially elicited against the V3 loop of the prototypic macrophage-tropic gp120 envelope protein ADA.     

Potent neutralization of primary HIV-1 isolates by antibodies directed against epitopes present in the V1/V2 domain of HIV-1 gp120

Although it is known that some human immune sera possess potent neutralizing activities for primary viruses, the identity of the target epitopes mediating this neutralization is unknown, and currently available immunogens have not been able to induce such activities. Using recombinant fusion glycoproteins expressing native V1/V2 domains of gp120 we have found that sera from a subset of HIV-1-infected humans contain antibodies that recognize broadly conserved V1/V2 epitopes. Such antibodies were isolated from one human serum by affinity chromatography on a column containing a V1/V2 fusion protein, and shown to efficiently neutralize several macrophage-tropic HIV-1 isolates. Rodents immunized with the purified V1/V2 fusion protein produced antibodies reactive with unrelated V1/V2 fusion proteins and with heterologous gp120s. V1/V2-specific immunoglobulins isolated from sera of these animals by affinity chromatography also possessed potent neutralization activity for several primary HIV-1 isolates. These results indicate that the V1/V2 domain of HIV-1 gp120 contains conserved epitopes that mediate potent neutralization of primary viruses, and suggest that subunit vaccines that efficiently induce such antibodies may provide protective humoral immunity against clinically relevant HIV-1 isolates.     

A deletion polymorphism due to Alu-Alu recombination in intron 2 of the retinoblastoma gene: association with human gliomas

In past years, much attention has been paid to the HIV-1 envelope (env) protein variable region 3 (V3), termed the principal neutralizing determinant. HIV-1 vaccines were often designed to target V3, and vaccine efficacy was often measured with V3-based assays. Thus, some disappointment resulted when volunteers in first clinical vaccine trials generated V3-specific antibodies that could not protect against V3-similar viruses. We describe an analysis of V1 and V2 sequence effects on antibody binding to V3 and non-V3 determinants. This study involved the preparation of seven full-length (gp160), chimeric env proteins in a vaccinia virus (VV) expression system. Chimeric proteins displayed different V1-V2 sequences but were otherwise identical. A panel of 12 monoclonal antibodies was then tested for binding activities toward the seven chimeras. Results showed that V1-V2 sequences affected antibody binding to env, both in V3 and non-V3 positions. These data demonstrate the enormous complexity of HIV-1 env protein conformation and antigenic determinants. Respect for the complexity of antibody-antigen interactions encourages the design of sophisticated immunoglobulin and protein cocktails for use in HIV-1 therapies and vaccines, respectively.     

Antibodies of symptomatic human immunodeficiency virus type 1-infected individuals are directed to the V3 domain of noninfectious and not of infectious virions present in autologous serum

The present study was designed to determine the antibody specificity for the human immunodeficiency virus type 1 (HIV-1) V3 domains of infectious and noninfectious virions present in the serum of AIDS patients. To accomplish this, HIV-1 was isolated in the presence of autologous antibodies from the serum samples of six AIDS patients in HIV-1-negative donor peripheral blood mononuclear cells by short-term cultivation. The isolated virus, defined as the infectious cell-free virus (iCFV), was characterized by sequence analysis of the proviral DNA coding for the third hypervariable (V3) region of the external glycoprotein gp120. This was carried out by amplifying and cloning the V3 region. In all six cases studied, 20 randomly selected V3 clones derived from the proviral DNA of the iCFV, 20 clones from patient cell-free virus, and 20 clones from cell-integrated virus were sequenced to study the distribution and frequency of the intrapatient virus population. The number of major virus variants in the six patients ranged from three to nine. The various V3 sequences found in the AIDS patients showed the typical amino acid pattern of the syncytium-inducing and non-syncytium-inducing viral phenotypes characteristic for the late stage of infection. However, only one patient-specific iCFV variant was detected within the 20 V3 clones analyzed per virus isolation. For the six patients a total of 34 V3-loop variants, either iCFV or non-iCFV, was observed. All 34 V3-loop sequences were expressed as glutathione-S-transferase fusion proteins (V3-GST). The autologous antibody response to the V3-GST fusion proteins was studied by Western immunoblot analysis. A strong antibody response to almost all non-iCFV V3-GST proteins was found in the sera of the six patients. In contrast, the autologous antibody response to the six iCFV V3 loops was undetectable (in four patients) or very faint (in two patients) compared with that to the non-iCFV V3 loops. Five of the six iCFV loops showed positively charged amino acids at positions strongly associated with the syncytium-inducing phenotype. These findings suggest that our in vitro isolation system selects for virions which are not recognized by V3-specific antibodies and are infectious both in vitro and in vivo.     

Recombinant human immunodeficiency Pr55gag virus-like particles presenting chimeric envelope glycoproteins induce cytotoxic T-cells and neutralizing antibodies

Very recently, we demonstrated that the replacement of the human immunodeficiency virus type-1 (HIV-1) gp41 transmembrane protein by an Epstein-Barr virus gp220/350-derived membrane anchor resulted in the incorporation of chimeric envelope (Env) oligomers into Pr55gag virus-like particles (VLPs), exceeding that of wild-type gp160 by a factor of 10. In this study, we examined the immunostimulatory properties of Pr55gag VLPs to both (i) chimeric HIV-1 gp120 external envelope proteins and (ii) full-length gp160 presented on the outer surface of the particles. Immunization studies carried out with VLPs presenting different derivatives of the chimeric and wild-type Env proteins elicited a consistent anti-Pr55gag as well as anti-Env antibody response in complete absence of additional adjuvants. In both cases, the immune sera exhibited an in vitro neutralizing activity against homologous HIV-1 infection in MT4 cells. Noteworthy, these VLPs were also capable of inducing a strong CD8+ cytotoxic T-cell (CTL) response in immunized BALB/c mice that was directed toward a known CTL epitope in the third variable domain V3 of the gp120 external glycoprotein. However, the induction of V3-loop-specific CTLs critically depended on the amounts of Env proteins that were presented by the Pr55gag VLPs. Moreover, the CD8+ CTL response was not significantly altered by adsorbing the VLPs to alum or by repeated booster immunizations. These results illustrate that Pr55gag VLPs provide a safe and effective means of enhancing neutralizing humoral responses to particle-entrapped gp120 proteins and are also capable of delivering these proteins to the MHC class I antigen processing and presentation pathway. Therefore, antigenically expanded Pr55gag VLPs represent an attractive approach in the design of vaccines for which specific stimulation of neutralizing antibodies and cytotoxic effector functions to complex glycoproteins is desired.     

Cross-reactions between the cytotoxic T-lymphocyte responses of human immunodeficiency virus-infected African and European patients

The great variability of protein sequences from human immunodeficiency virus (HIV) type 1 (HIV-1) isolates represents a major obstacle to the development of an effective vaccine against this virus. The surface protein (Env), which is the predominant target of neutralizing antibodies, is particularly variable. Here we examine the impact of variability among different HIV-1 subtypes (clades) on cytotoxic T-lymphocyte (CTL) activities, the other major component of the antiviral immune response. CTLs are produced not only against Env but also against other structural proteins, as well as some regulatory proteins. The genetic subtypes of HIV-1 were determined for Env and Gag from several patients infected either in France or in Africa. The cross-reactivities of the CTLs were tested with target cells expressing selected proteins from HIV-1 isolates of clade A or B or from HIV type 2 isolates. All African patients were infected with viruses belonging to clade A for Env and for Gag, except for one patient who was infected with a clade A Env-clade G Gag recombinant virus. All patients infected in France were infected with clade B viruses. The CTL responses obtained from all the African and all the French individuals tested showed frequent cross-reactions with proteins of the heterologous clade. Epitopes conserved between the viruses of clades A and B appeared especially frequent in Gag p24, Gag p18, integrase, and the central region of Nef. Cross-reactivity also existed among Gag epitopes of clades A, B, and G, as shown by the results for the patient infected with the clade A Env-clade G Gag recombinant virus. These results show that CTLs raised against viral antigens from different clades are able to cross-react, emphasizing the possibility of obtaining cross-immunizations for this part of the immune response in vaccinated individuals.     

Factors affecting the immunogenicity and potency of tetanus toxoid: implications for the elimination of neonatal and non-neonatal tetanus as public health problems

The neutralizing activity of anti-V3 monoclonal antibodies (MAbs) and anti-HIV-1 immune sera was tested against HIV-1 laboratory strains and African primary isolates. Neutralization was investigated in Phytohaemagglutinin (PHA)-stimulated peripheral blood mononuclear cell (PBMC) cultures by means of two distinct viral titer reduction assays. In these assays, virus was detected by means of either p24 antigen measurement using ELISA or HIV provirus synthesis using PCR, respectively. Anti-V3 MAbs and anti-HIV-1 immune sera neutralized efficiently the homologous laboratory HIV-1 strains used for eliciting immune response but showed no neutralizing activity against most primary isolates. The two neutralization assays used provided similar results. However, a PCR-based assay circumvented the limitations due to low levels of virus replication. The mechanism of resistance of the primary isolates to neutralizing antibodies was complex and was not simply predicted by partial sequence determination of the epitopes. This points out the need for reliable neutralization assays of HIV-1 primary isolates in order to evaluate the role of humoral immunity during HIV-1 infection and for future vaccine strategies.     

Induction of neutralizing antibodies to T-cell line-adapted and primary human immunodeficiency virus type 1 isolates with a prime-boost vaccine regimen in chimpanzees

Five chimpanzees were immunized by administration of one or more intranasal priming doses of one to three recombinant adenoviruses containing a gp160 insert from human immunodeficiency virus type 1 (HIV-1) MN (HIV-1MN) followed by one or more boosts of recombinant HIV-1SF2 gp120 delivered intramuscularly with MF59 adjuvant. This regimen resulted in humoral immune responses in three of five animals. Humoral responses included immunochemically active anti-H1V-1 antibodies (Abs) directed to recombinant gp120 and neutralizing Abs reactive with T-cell-line-adapted HIV-1MN and HIV-1SF2. In addition, neutralizing activity was detected to the two homologous primary isolates and to two of three heterologous primary isolates which, like the immunizing strains, can use CXCR4 as a coreceptor for infection. The three animals with detectable neutralizing Abs and a fourth exhibiting the best cytotoxic T-lymphocyte response were protected from a low-dose intravenous challenge with a cell-free HIV-1SF2 primary isolate administered 4 weeks after the last boost. Animals were rested for 46 weeks and then rechallenged, without a boost, with an eightfold-higher challenge dose of HIV-1SF2. The three animals with persistent neutralizing Abs were again protected. These data show that a strong, long-lived protective Ab response can be induced with a prime-boost regimen in chimpanzees. The data suggest that in chimpanzees, the presence of neutralizing Abs correlates with protection for animals challenged intravenously with a high dose of a homologous strain of HIV-1, and they demonstrate for the first time the induction of neutralizing Abs to homologous and heterologous primary isolates.     

Variability analysis of HIV-1 gp120 V3 region: IV. Distribution functions for intra- and inter-subtype amino acid hamming distances

Distribution functions for intra- and inter- HIV-1 V3-loop subtypes amino acid Hamming distances were calculated (850 V3-loop sequences from the Los Alamos HIV-1 Database (1996) were used). These functions have pronounced bell-like shape. Such shapes of the histograms for HIV-1 V3 intra- and inter-subtype distriutions are discussed to confirm the applicability of different hierarchical cluster procedures for HIV-1 V3 classification. Two-mode distribution for the subtype E could sertificate that this subtype includes two thinner taxons.     

beta-chemokines and neutralizing antibody titers correlate with sterilizing immunity generated in HIV-1 vaccinated macaques

One of the obstacles to AIDS vaccine development is the variability of HIV-1 within individuals and within infected populations, enabling viral escape from highly specific vaccine induced immune responses. An understanding of the different immune mechanisms capable of inhibiting HIV infection may be of benefit in the eventual design of vaccines effective against HIV-1 variants. To study this we first compared the immune responses induced in Rhesus monkeys by using two different immunization strategies based on the same vaccine strain of HIV-1. We then utilized a chimeric simian/HIV that expressed the envelope of a dual tropic HIV-1 escape variant isolated from a later time point from the same patient from which the vaccine strain was isolated. Upon challenge, one vaccine group was completely protected from infection, whereas all of the other vaccinees and controls became infected. Protected macaques developed highest titers of heterologous neutralizing antibodies, and consistently elevated HIV-1-specific T helper responses. Furthermore, only protected animals had markedly increased concentrations of RANTES, macrophage inflammatory proteins 1alpha and 1beta produced by circulating CD8(+) T cells. These results suggest that vaccine strategies that induce multiple effector mechanisms in concert with beta-chemokines may be desired in the generation of protective immune responses by HIV-1 vaccines.     

Identification of the B cell superantigen-binding site of HIV-1 gp120

Previous studies showed that the gp120 envelope protein of HIV-1 is able to crosslink membrane IgM on normal human B cells and to induce their activation in a V(H)3 immunoglobulin gene-family-specific manner. Because this V(H) gene family is the largest in the human repertoire, this superantigen (SAg) property is thought to have deleterious consequences for the host, including a progressive decline of B cells with progression of the HIV-1-induced disease. Here, we have identified the sequence motifs on gp120 involved in SAg binding to normal Igs. We show that this SAg-binding activity is present in gp120s from highly divergent isolates of HIV-1 belonging to clades derived from various geographical origins, and that carbohydrate residues are not essential for its expression. The SAg-binding site is formed by protein sequences from two regions of the gp120 molecule. The core motif is a discontinuous epitope spanning the V4 variable domain and the amino-terminal region flanking the C4 constant domain. The most critical residues appear to be Leu395-Asp397 and Ile425-Gln427. Residues from the C2 constant domain (positions 252-272) also seem to play an accessory role in SAg binding of gp120 to normal human Igs. These findings are important in the design of a successful gp120-based vaccine against HIV-1.     

Study of the V3 loop as a target epitope for antibodies involved in the neutralization of primary isolates versus T-cell-line-adapted strains of human immunodeficiency virus type 1

Previous studies characterized the third variable (V3) loop of the envelope gp120 as the principal neutralizing determinant for laboratory T-cell-line-adapted (TCLA) strains of human immunodeficiency virus type 1 (HIV-1). However, primary viruses isolated from infected individuals are more refractory to neutralization than TCLA strains, suggesting that qualitatively different neutralizing antibodies may be involved. In this study, we investigated whether the V3 loop constitutes a linear target epitope for antibodies neutralizing primary isolates. By using peptides representative of the V3 regions of various primary isolates, an early, relatively specific and persistent antibody response was detected in sera from HIV-infected patients. To assess the relationship between these antibodies and neutralization, the same peptides were used in competition and depletion experiments. Addition of homologous V3 peptides led to a competitive inhibition in the neutralization of the TCLA strain HIVMN/MT-4 but had no effect on the neutralization of the autologous primary isolate. Similarly, the removal of antibodies that bind to linear V3 epitopes resulted in a loss of HIVMN/MT-4 neutralization, whereas no decrease in the autologous neutralization was measured. The different roles of V3-specific antibodies according to the virus considered were thereby brought to light. This confirmed the involvement of V3 antibodies in the neutralization of a TCLA strain but emphasized a more pronounced contribution of either conformational epitopes or epitopes outside the V3 loop as targets for antibodies neutralizing primary HIV-1 isolates. This result underlines the need to focus on new vaccinal immunogens with epitopes able to induce broadly reactive and efficient antibodies that neutralize a wide range of primary HIV-1 isolates.     

Lack of correlation between V3-loop peptide enzyme immunoassay serologic subtyping and genetic sequencing

OBJECTIVE: To compare the performance of V3-loop peptide enzyme immunoassay (PEIA) methodologies from four different laboratories for subtyping HIV-1, and to determine the causes for the lack of correlation between V3-loop PEIA serotyping and subtyping by sequencing. MATERIALS AND METHODS: Synthetic peptides derived from the amino-acid consensus sequences of the V3-loop of group M strains representing genetic subtypes A-F as well as reference strains were evaluated in PEIA by four different laboratories for their ability to accurately determine the subtype in a panel of 85 sera obtained from persons infected with known HIV-1 subtypes (28 subtype A, 34 subtype B, four subtype C, 10 subtype D, seven subtype F, one each of subtype H and G). Furthermore, the V3 loop of the corresponding virus was compared with the V3 loop of the peptides used in PEIA. RESULTS: The correlation between HIV-1 subtyping by sequencing and V3-loop PEIA from the different laboratories varied considerably for the different HIV-1 subtypes: subtype A (46-68%), B (38-85%), C (75-100%), D (29-50%), and F (17-57%). A 70% agreement between PEIA and sequencing subtypes was observed for samples with the concordant presence of the same octameric sequences in the V3 loop of the virus and the V3 loop of the peptide used in PEIA; however, only 42% of specimens with different V3-loop octameric viral and peptide sequences yielded concordant results in V3-loop serotyping and genetic subtyping. CONCLUSION: Our results indicate that V3-loop PEIA methodologies used in different laboratories correlate poorly with genetic subtyping, and that their accuracy to predict HIV-1 subtypes in sera of Belgian individuals infected with different HIV-1 subtypes (A, B, C, D, F, G and H) vary considerably. The poor correlation between serotyping and genetic subtyping was partly due to the simultaneous occurrence of subtype-specific octameric sequences at the tip of the V3 loop of viruses belonging to different genetic subtypes.     

Immune response to baculovirus expressed protein fragment amino acids 190-289 of respiratory syncytial virus (RSV) fusion protein

At least two neutralizing epitopes have been identified in the amino acid (aa) sequence 190-289 of the RSV fusion protein. The authors expressed this region in insect cells (bF190-289) and compared the immune response to bF190-289 with that induced by baculovirus expressed full-length fusion protein (bF). As with bF, mice primed with bF190-289 produced exclusively antibodies of IgG1 isotype, generated neutralizing antibodies, reduced significantly the virus titer (about a half log10 reduction) after RSV challenge and induced a Helper T (Th) 2 cell response in mediastinal lymph node cells (MLNC) restimulated in vitro. Thus, the aa sequence 190-289 represents a major immunogenic region of the RSV fusion protein.     

Neutralizing antibodies from the sera of human immunodeficiency virus type 1-infected individuals bind to monomeric gp120 and oligomeric gp140

Antibodies that neutralize primary isolates of human immunodeficiency virus type 1 (HIV-1) appear during HIV-1 infection but are difficult to elicit by immunization with current vaccine products comprised of monomeric forms of HIV-1 envelope glycoprotein gp120. The limited neutralizing antibody response generated by gp120 vaccine products could be due to the absence or inaccessibility of the relevant epitopes. To determine whether neutralizing antibodies from HIV-1-infected patients bind to epitopes accessible on monomeric gp120 and/or oligomeric gp140 (ogp140), purified total immunoglobulin from the sera of two HIV-1-infected patients as well as pooled HIV immune globulin were selectively depleted of antibodies which bound to immobilized gp120 or ogp140. After passage of each immunoglobulin preparation through the respective columns, antibody titers against gp120 and ogp140 were specifically reduced at least 128-fold. The gp120- and gp140-depleted antibody fraction from each serum displayed reduced neutralization activity against three primary and two T-cell line-adapted (TCLA) HIV-1 isolates. Significant residual neutralizing activity, however, persisted in the depleted sera, indicating additional neutralizing antibody specificities. gp120- and ogp140-specific antibodies eluted from each column neutralized both primary and TCLA viruses. These data demonstrate the presence and accessibility of epitopes on both monomeric gp120 and ogp140 that are specific for antibodies that are capable of neutralizing primary isolates of HIV-1. Thus, the difficulties associated with eliciting neutralizing antibodies by using current monomeric gp120 subunit vaccines may be related less to improper protein structure and more to ineffective immunogen formulation and/or presentation.     

Expression and immune response to foreign epitopes in bacteria. Perspectives for live vaccine development

We previously developed a general procedure which allows the genetic coupling of a chosen foreign linear epitope in different regions of a carrier protein. By using as carriers, two bacterial envelope proteins, the LamB and MalE proteins of E. coli K12, we were able to express the same epitope in different sites of the two proteins and in different compartments of the bacteria. This allowed us to analyze the influence of the localization in E. coli cells of a foreign B-cell epitope on the induction of specific antibody responses, and the role of the molecular environment on the immunological properties of foreign B- or T-cell epitopes, using either purified hybrid proteins or live recombinant bacteria. Several LamB and MalE hybrid proteins were expressed in the aroA attenuated strain of S. typhimurium, SL3261. Immunizations of mice with live recombinant bacteria by the intravenous route showed that it was possible to induce humoral responses against inserted foreign sequences. In order to improve the in vivo stability of the plasmids carrying the different contructions, and to increase the amounts of recombinant LamB and MalE hybrid proteins expressed in vivo, the LamB and malE genes were placed under the control of the anaerobically inducible pnirBpromoter control. The genetic factors susceptible of influencing the immune response to recombinant Salmonella in mice were also studied.     

The human interleukin-6 receptor alpha chain gene is localized on chromosome 1 band q21

An overview of efforts to induce neutralizing antibodies in order to develop an effective vaccine against AIDS is presented. The principal Neutralizing Determinant (PND) on the HIV-1 envelope is described. PND variability and the induction of neutralizing antibodies by synthetic peptides representing PND are discussed. The use of a cocktail of different peptides representing the PND sequences of the majority of HIV-1 isolates, as well as the construction of hybrid immunogens containing PND of several viral isolates, could overcome the problems related to PND variability. A different approach based on the possibility of inducing a type of intracellular immunity is also discussed: a cellular clone (F12) obtained in our laboratory from Hut-78 cells infected with supernatant of cultured lymphocytes from an HIV-infected patient, does not release viral particles despite the presence of a full-length HIV-1 provirus. Moreover, F12 cells are fully resistant against superinfection with any HIV-1 or HIV-2 isolates. We are now attempting to reproduce the homologous viral interference by transferring the F12/HIV genome of the clone into HIV-susceptible cells in order to render these cells resistant to HIV infection.     

Neutralization of HIV-1 by secretory IgA induced by oral immunization with a new macromolecular multicomponent peptide vaccine candidate

Control of pandemic infection of human immunodeficiency virus type 1 (HIV-1) requires some means of developing mucosal immunity against HIV-1 because sexual transmission of the virus occurs mainly through the mucosal tissues. However, there is no evidence as yet that the secretory immunoglobulin A (IgA) antibody induced by immunization with antigens in experimental animals can neutralize HIV-1. We demonstrate here that oral immunization with a new macromolecular peptide antigen and cholera toxin (CT) induces a high titre (1:2") of gut-associated and secretory IgA antibody to HIV-1. Using three different neutralizing assays, we clearly demonstrate that this secretory IgA antibody is able to neutralize HIV-1IIIB, HIV-1SF2 and HIV-1MN. Our new approach may prove to be important in the development of a mucosal vaccine that will provide protection of mucosal surfaces against HIV-1.     

Characterization of simian-human immunodeficiency virus envelope glycoprotein epitopes recognized by neutralizing antibodies from infected monkeys

We characterized human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein epitopes recognized by neutralizing antibodies from monkeys recently infected by molecularly cloned simian-human immunodeficiency virus (SHIV) variants. The early neutralizing antibody response in each infected animal was directed mainly against a single epitope. This primary neutralizing epitope, however, differed among individual monkeys infected by identical viruses. Two such neutralization epitopes were determined by sequences in the V2 and V3 loops of the gp120 envelope glycoprotein, while a third neutralization epitope, apparently discontinuous, was determined by both V2 and V3 sequences. These results indicate that the early neutralizing antibody response in SHIV-infected monkeys is monospecific and directed against epitopes composed of the gp120 V2 and V3 variable loops.